Human activation-induced deaminase lacks strong replicative strand bias or preference for cytosines in hairpin loops

Abstract

Activation-induced deaminase (AID) is a DNA-cytosine deaminase that mediates maturation of antibodies through somatic hypermutation and class-switch recombination. While it causes mutations in immunoglobulin heavy and light chain genes and strand breaks in the switch regions of the immunoglobulin heavy chain gene, it largely avoids causing such damage in the rest of the genome. To help understand targeting by human AID, we expressed it in repair-deficient Escherichia coli and mapped the created uracils in the genomic DNA using uracil pull-down and sequencing, UPD-seq. We found that both AID and the human APOBEC3A preferentially target tRNA genes and transcription start sites, but do not show preference for highly transcribed genes. Unlike A3A, AID did not show a strong replicative strand bias or a preference for hairpin loops. Overlapping uracilation peaks between these enzymes contained binding sites for a protein, FIS, that helps create topological domains in the E. coli genome. To confirm whether these findings were relevant to B cells, we examined mutations from lymphoma and leukemia genomes within AID-preferred sequences. These mutations also lacked replicative strand bias or a hairpin loop preference. We propose here a model for how AID avoids causing mutations in the single-stranded DNA found within replication forks.

An APOBEC3 Mutational Signature in the Genomes of Human-Infecting Orthopoxviruses

“…The same authors also reported transcriptional asymmetry of APOBEC3 mutation signatures, a feature we do not observe in hMPXV1 genes. However, both the association with transcription levels and transcription asymmetry are still controversial findings in human cancer genomes ( 10 – 12 , 36 , 37 ). Thus, experimental analyses will be required to determine the molecular mechanisms of APOBEC3-mediated editing of hMPXV1 genomes and the role of transcription.…”

Distinguishing preferences of human APOBEC3A and APOBEC3B for cytosines in hairpin loops, and reflection of these preferences in APOBEC-signature cancer genome mutations

“…1B ). The tendency of AID/APOBEC enzymes of preferentially deaminating cytosines near TSS and within tRNA genes in E. coli , yeast and human tumors has been noted previously 44 , 46 , 49 , 50 .

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“…This analysis confirmed that A3A targets hairpin loops 46 and further showed that cytosines in VpC dinucleotides (V is A, C or G) are much better targets for A3A when they appear in hairpin loops than in linear DNA 41 . UPD-seq also showed that AID lacks a strong preference for LGST in replication forks and does not prefer cytosines in hairpins 44 . Normal B lymphocytes undergoing somatic hypermutation and class-switch recombination express AID during the G1 phase of the cell cycle 47 , and hence these results were interpreted to mean that ssDNA in replication forks is effectively protected by RPA from the action of any AID that may leak into the S phase 44 .…”

“…UPD-seq also showed that AID lacks a strong preference for LGST in replication forks and does not prefer cytosines in hairpins 44 . Normal B lymphocytes undergoing somatic hypermutation and class-switch recombination express AID during the G1 phase of the cell cycle 47 , and hence these results were interpreted to mean that ssDNA in replication forks is effectively protected by RPA from the action of any AID that may leak into the S phase 44 .…”

“…A related study 43 , showed that the ssDNA-binding protein, RPA, can affect the activity of A3A, generally inhibiting it. This has led to the suggestion that there is a competition between RPA and the APOBECs for binding to ssDNA, especially at replication forks 41 , 44 , 45 .…”